This twenty-year old grant has as its theme the study of morphogenesis, with particular emphasis on correlation of cytology with biochemistry and with experimental studies of tissue interaction and cell differentiation. In the five-year period for which renewal is sought, we will continue to concentrate on the embryonic extracellular matrix, its role in morphogenesis, its structural relations to cell surfaces, and the interrelations among its constituent molecules. The work contemplated can be outlined as follows: I. Localization of Morphogenetically Significant Molecules in Embryonic Tissues. The time is ripe to exploit immunohistochemistry and specific strains at the ultrastructural level (TEM and SEM) to localize collagens, glycosaminoglycans, and fibronectins. The developing cornea, embryonic trunk limb and area vasculosa will be studied along these lines. We seek innovative solutions to problems confronting electron microscopic immunohistochemistry: we will develop directly labeled monoclonal antibodies, alternatives to ferritin-labels, double-viewing by TEM and SEM, and double antibody-labeling. We will combine this approach with freeze-cleave and deep-etch to analyze the relation of cell surface collagen to the cortical cytoskeleton. Immunohistochemistry and polycationic stains will be used in conjunction with TEM and SEM to identify nonstriated fibrils and glycosaminoglycan granules in the ECM and to map the distribution of fibronectin in the early chick and rat embryo. II. Further Experiments on the Role of Extracellular Molecules in Morphogenesis. We will study the disruption in the organization of the corneal epithelial basal surface that takes place when it is isolated by EDTA; the effect of soluble and insoluble collagens on the restoration of epithelial differentiation will be analyzed in some detail. Finally, pilot experiments will be started on the effect of epithelial ECM and demineralized bone matrix on fibroblast differentiation in vitro and of injected glycosaminoglycans on amphibian limb regeneration in situ.